The present invention relates to the field of plant molecular biology, more particularly Jatropha microsomal ω6 oleate desaturases. The present invention also relates to Jatropha plants or plants of other oil crops having seeds with altered ratios of monosaturated and polyunsaturated fats. In particular, the present invention relates to Jatropha plants or plants of other oil crops where the plants exhibit elevated levels of oleic acid.
The publications and other materials used herein to illuminate the background of the invention, and in particular, cases to provide additional details respecting the practice, are incorporated by reference, and for convenience are referenced in the following text by author and date and are listed alphabetically by author in the appended bibliography.
Plant oils have many kinds of diverse applications. Novel vegetable oil compositions and improved approaches to obtain oil compositions, from biosynthetic or natural plant sources, are needed. Depending upon the intended oil use, various different fatty acid compositions are desired. Plants, especially species which synthesize large amounts of oils in seeds, are an important source of oils both for edible and industrial uses (Lu et al.; Durrett et al., 2008).
One major usage for plant oil is for food. Plant oils are mostly composed of five common fatty acids, namely palmitic acid (16:0), stearic acid (18:0), oleic acid (18:1), linoleic acid (18:2) and linolenic acid (18:3) (Durrett et al., 2008). Oleic acid is a monounsaturated omega-9 and 18 carbon fatty acid found in various vegetable oils. It is considered one of the healthier sources of oil and fat in food resources for human and animal. Diets in which oil consumption are high in oleic acid have been proven to downregulate overall levels of chronic human diseases such as cholesterol, arteriosclerosis and cardiovascular disease. Specifically, oleic acid has been shown to raise levels of high-density lipoproteins (HDLs) known as “good cholesterol”, while lowering low-density lipoproteins (LDLs) also known as the “bad” cholesterol. Thus, the development of new and inexpensive sources of foods comprising healthier forms of fatty acid is desirable.
One emerging purpose for oil is to serve as feedstock of renewable bioenergy in the form of biodiesel. The demand for use biodiesel, mainly comes from vegetable oil, has soared along with government subsidies and mandates for the alternative fuel. Because there are various fatty acid composition of each types, the fuel properties of biodiesel derived from a mixture of fatty acids are dependent on that composition. Compared with conventional diesel, there are some negative factors of fatty acid profile should be optimized by traditional breeding or genetic engineering to optimize biodiesel fuel characteristics. Various studies suggest that biodiesel with high levels of methyl oleate will have excellent, characteristics with regard to ignition quality, NOx emissions and fuel stability. For example, while unsaturation tends to reduce the cetane number of biodiesel, that of methyl oleate is higher than the minimal biodiesel standard. Additionally, it has been estimated that biodiesel fuels with an average of 1.5 double bonds per molecule will emit an equivalent amount of NOx compared with conventional diesel, thus a fuel high in oleates should not result in higher NOx emissions. Finally, given that polyunsaturated fatty acids have a major effect on the auto-oxidation of biodiesel, high oleic acid with reduced polyunsaturated fatty acid content will improve the stability of the fuel (Durrett et al., 2008).
Soybean lines with high levels of oleic acid and low levels of saturated and polyunsaturated fatty acids have been developed using a transgenic strategy that results in down-regulation of one single gene fatty acid desaturase 2 (FAD2). Consistent with predictions, biodiesel synthesized from these high-oleic soybeans demonstrated improved fuel characteristics with regard to cold-temperature flow properties and NOx emissions (Tat et al., 2007; Graef et al., 2009).
During the last several years, many countries have begun to target biofuel research as a national priority and implement compulsory blending of fossil fuel with biofuel. The increasing demand for biofuel, however, is exerting more pressure on food production because of the competition between fuel crops and food crops for arable land. One way to ease this competition is to use marginal land for bio-energy production (Carroll and Somerville, 2008).
Jatropha curcas, a small woody plant belonging to Euphorbiaceae, is a non-food crop mainly grown in the tropical and subtropical regions. This plant possesses several properties rendering it suitable for biodiesel production, such as its rapid growth, ease of propagation, short gestation period, low seed cost, high oil content, wide adaptability, and drought tolerance (Jones N, 1991; Fairless, 2007). Furthermore, Jatropha may yield more than four times as much fuel per hectare as soybean, and more than ten times that of maize (corn) (http://en.wikipedia.org/wiki/Jatropha_oil). Especially important is that Jatropha can thrive on degraded soil (Fairless, 2007) making it an attractive crop for biodiesel feedstock since it can be planted on a large-scale on marginal land unsuitable for food crops.
Plants synthesize fatty acids via a common metabolic pathway known as the fatty acid synthase (FAS) pathway. Beta-ketoacyl-ACP (acyl carrier protein moiety) synthases are important rate-limiting enzymes in the FAS of plant cells and exist in several versions. Beta-ketoacyl-ACP synthase I catalyzes chain elongation to palmitoyl-ACP (C16:0), whereas Beta-ketoacyl-ACP synthase II catalyzes chain elongation to stearoyl-ACP (C18:0). Beta-ketoacyl-ACP synthase IV is a variant of Beta-ketoacyl-ACP synthase II, and can also catalyze chain elongation to 18:0-ACP. In soybeans, the major products of FAS are 16:0-ACP and 18:0-ACP. The desaturation of 18:0-ACP to form 18:1-ACP is catalyzed by a plastid-localized soluble delta-9 desaturase (also referred to as “stearoyl-ACP desaturase”).
The products of the plastidial FAS and delta-9 desaturase, 16:0-ACP, 18:0-ACP, and 18:1-ACP, are hydrolyzed by specific thioesterases. Plant thioesterases can be classified into two gene families based on sequence homology and substrate preference. The first family, FATA, includes long chain acyl-ACP thioesterases having activity primarily on 18:1-ACP. Enzymes of the second family, FATB, commonly utilize 16:0-ACP (palmitoyl-ACP), 18:0-ACP (stearoyl-ACP), and 18:1-ACP (oleoyl-ACP). Such thioesterases have an important role in determining chain length during de novo fatty acid biosynthesis in plants, and thus these enzymes are useful in the provision of various modifications of fatty acyl compositions, particularly with respect to the relative proportions of various fatty acyl groups that are present in seed storage oils.
The products of the FATA and FATB reactions, the free fatty acids, leave the plastids and are converted to their respective acyl-CoA esters. Acyl-CoAs are substrates for the lipid-biosynthesis pathway (Kennedy Pathway), which is located in the endoplasmic reticulum (ER). This pathway is responsible for membrane lipid formation as well as the biosynthesis of triacylglycerols, which constitute the seed oil. In the ER there are additional membrane-bound desaturases, which can further desaturate 18:1 to polyunsaturated fatty acids.
Various technologies for generating mid to high oleic acid levels in soybean plants are known. For example, U.S. Patent Publication No. 2007/0214516 discloses a method for obtaining soybean plants that have moderately increased levels of oleic acid.